The Paper We’ve Been Waiting For? CRISPR/Cas9 in Human Embryonic Stem Cells

Well it seems it has been done, the paper showing off the stuff science fiction has come out. CRISPR/Cas9 was used in human embryonic stem cells by the Meissner group at Harvard featuring J. Keith Joung and John Rinn.

“Targeted disruption of DNMT1, DNMT3A and DNMT3B in human embryonic stem cells”

Nature Genetics

Abstract: “DNA methylation is a key epigenetic modification involved in regulating gene expression and maintaining genomic integrity. Here we inactivated all three catalytically active DNA methyltransferases (DNMTs) in human embryonic stem cells (ESCs) using CRISPR/Cas9 genome editing to further investigate the roles and genomic targets of these enzymes. Disruption of DNMT3Aor DNMT3B individually as well as of both enzymes in tandem results in viable, pluripotent cell lines with distinct effects on the DNA methylation landscape, as assessed by whole-genome bisulfite sequencing. Surprisingly, in contrast to findings in mouse, deletion of DNMT1 resulted in rapid cell death in human ESCs. To overcome this immediate lethality, we generated a doxycycline-responsive tTA-DNMT1* rescue line and readily obtained homozygous DNMT1-mutant lines. However, doxycycline-mediated repression of exogenous DNMT1* initiates rapid, global loss of DNA methylation, followed by extensive cell death. Our data provide a comprehensive characterization of DNMT-mutant ESCs, including single-base genome-wide maps of the targets of these enzymes.”

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